知母甾体皂苷抗血小板活性作用机制的研究
|
摘要
血小板是血液中由骨髓巨核细胞脱落下无核的小块胞质,在机体初期止血中起着重要作用。此外在病理条件下,血小板也与一些心脑血管疾病密切相关。越来越多的证据显示,异常的血小板激活是动脉粥样硬化形成中的一个重要因素。目前,心脑血管疾病仍然是导致人类死亡的主要原因之一,抗血小板治疗成为治疗此类疾病的有效策略。许多抗血小板药物如阿司匹林(环氧合酶抑制剂)、氯吡咯雷和噻氯匹啶(ADP受体阻断剂)对心血管疾病有很好的疗效,然而,在临床上也观察到这些药物存在药物抵抗、中性粒细胞减少、出血、血小板减少等副作用。因此,研发更加安全有效的抗血小板药物是治疗和预防动脉栓塞性疾病有效的途径。
甾体皂苷是一类重要的中药活性成分,在许多植物中都含有甾体皂苷成分。它具有广泛的药理作用和重要的生物活性,例如防治心血管疾病、抗肿瘤、抗真菌等作用。甾体皂苷的基本骨架是由含27个碳的苷元结构与一个或多个糖链连接构成的,主要分为含一个糖链(主要在C3-位)的螺甾皂苷(spirostanol saponins)和含2个糖链(主要在C 3-及C26-位)的呋甾皂苷(furostanol saponins)。甾体皂苷具有较高的医药价值,已有报道多种甾体皂苷具有明确的抗血小板活性作用,但目前对甾体皂苷抗血小板作用机制还了解很少。本研究的目的就是揭示甾体皂苷抗血小板活性的作用机制。
我们首先筛选了六个从知母中提取和合成的单体化合物,发现知母皂苷AⅢ抑制血小板聚集的作用最强。进一步构效关系分析显示,苷元结构和C-3位糖链组成和连接方式与甾体皂苷抑制血小板聚集活性密切相关。体内研究结果表明知母皂苷AⅢ延长小鼠尾静脉出血时间,保护致死性的血栓栓塞。这些结果提示知母皂苷AⅢ是研究甾体皂苷抗血小板活性的理想小分子探针。
我们进一步研究知母皂苷AⅢ抗血小板活性的作用特点。血小板激活包括变形、聚集、释放等反应。血小板抑制剂可以抑制血小板的激活反应。在我们研究中,知母皂苷AⅢ抑制了血小板聚集、变形、释放、αIIbβ3激活、蛋白磷酸化等激活反应。同时,知母皂苷AⅢ也表现出其抗血小板活性的自身特点,知母皂苷AⅢ虽然剂量依赖性地抑制U46619(TXA2受体激动剂)、AA、胶原和ADP诱导的血小板聚集,但对各诱导剂的敏感性不同,对U46619和AA的作用最强,这提示知母皂苷AⅢ可能阻断TXA2生成或直接作用TXA2通路上;观察到知母皂苷AⅢ本身也会引起血小板变形,而PGE1则没有观察到这一现象。PGE1与血小板膜上IP受体结合,激活Gs通路,升高细胞内cAMP。升高的cAMP能阻断多条血小板信号通路激活而抑制血小板的激活反应。然而,在我们的实验中并未观察到知母皂苷AⅢ升高血小板cAMP的作用。知母皂苷AⅢ与PGE1相似,是一个有效的血小板抑制剂,但其发挥作用并不是通过升高cAMP。
PKC的激活与血小板激活反应有关,主要调节血小板的聚集和释放。PMA激活PKC诱导血小板聚集和释放,然而,知母皂苷AⅢ并没有抑制PMA诱导的血小板变形和释放,这提示知母皂苷AⅢ可能抑制PKC上游信号通路。凝血酶与血小板膜G蛋白偶联受体结合,激活磷脂酶C,产生DAG,导致PKC的激活,引起血小板的释放反应。凝血酶激活血小板仅在低浓度作用时依赖TXA2的生成,而高浓度时不依赖TXA2的生成。使用高浓度凝血酶诱导血小板聚集,知母皂苷AⅢ不能抑制高浓度凝血酶诱导的血小板聚集,而选择性抑制TXA2受体激动剂U46619介导的血小板聚集和释放。我们的实验证实了U46619、凝血酶和PMA都能诱导血小板ERK的磷酸化,知母皂苷AⅢ能有效地抑制U46619诱导ERK1/2磷酸化,而对不依赖TXA2受体介导的凝血酶和PMA诱导的ERK1/2磷酸化没有作用,这表明知母皂苷AⅢ能阻断TXA2受体介导的信号通路的激活。血小板激活生成TXA2分泌后与血小板膜表面TXA2受体结合,而TXA2受体主要偶联在Gq和G12/13两条通路上,Gq通路介导的PLCβ激活在激动剂诱导的血小板聚集和释放中有至关重要的作用;G12/13介导的Rho/Rho激酶途径主要与血小板的变形有关。我们研究表明,知母皂苷AⅢ选择性阻断TXA2受体介导的Gq通路的激活,而不影响G12/13通路。此外,磷脂酶C抑制剂能抑制知母皂苷AⅢ诱导的血小板变形,知母皂苷AⅢ诱导的变形很可能由于部分激活了Gq通路。最后,我们发现知母皂苷AⅢ能协同TXA2受体拮抗剂SQ29548共同抑制U46619诱导的血小板聚集。
总的来说,我们选择知母皂苷AⅢ作为理想的小分子探针研究甾体皂苷抗血小板活性的作用机制,并在研究信号转导通路的基础上,推测TXA2受体可能是知母皂苷AⅢ抗血小板活性作用的候选靶点。
Platelets, anucleated cells that originate from bone marrow megakaryocytes, are essential for primary hemostasis in the blood. In addition, a role for platelets in pathogenesis of a number of cardiovascular events is recognized. Increasing evidence indicates that enhanced platelet activation plays a critical role in initiation and development of atherothrombotic diseases. Accordingly, to prevent and treat thrombosis and vascular diseases, antiplatelet therapy may be a potential strategy. Many antiplatelet agents such as aspirin (an inhibitor of cyclooxygenase (COX)), clopidogrel, and ticlopidine (adenosine diphosphate receptor antagonists acting at the purinergic receptors of P2Y), have been reported to be beneficial to patients suffering from cardiovascular disorders. However, some drawbacks of these drugs such as aspirin resistance, the risk of neutropenia, bleeding, and thrombocytopenia, were occasionally observed. Therefore, development of more safe and effective antiplatelet agents may be a promising approach for prevention and treatment of atherothrombosis.
Steroidal saponins are widely distributed in plants and have many pharmacologic actions and biological activities, such as antiplatelet activity, antitumor activity, and antidementia activity. They are an important class of natural products which is composed of a C-27 aglycone moiety and sugar chains of one or more monosaccharides. These compounds are classified as spirostanol glycosides with a sugar chain at C3 position, and furostanol saponins with two sugar chains at both C3 and C26 positions respectively. They have considerable potential as pharmaceutical and/or nutraceutical agents in natural or synthetic form. Steroidal glycosides, from a variety of sources, have been reported to have platelet inhibitory or aggregative activity, such as sarsasapogenin glycosides extracted from the rhizome of Anemarrhenae asphodeloides Bge (Liliaceae), has shown strong anti-platelet activity. But until now, little is known about molecular mechanism for the anti-platelet activity .Our aim is to determine the mechanism of steroidal saponins of anti-platelet activity.
We first screened 6 natural steroidal saponins isolated from the root of Anemarrhenae asphodeloides Bge (Liliaceae) for their effects on rat platelet aggregation. We found that Timosaponin AⅢexhibited stronger inhibitory effects on platelet aggregation than other compounds, Further structure–activity assay revealed that the activity of Timosaponin AⅢin inhibiting platelet aggregation was not only attributed in part to the number, the length and the type of sugar side chains attached by a glycoside at C-3, but also related with the aglycon part. In the in vivo study, Timosaponin AⅢprolonged mouse tail bleeding time using bleeding time model and protected mice against fatal thrombosis in a concentration dependent manner. These results indicated that Timosaponin AⅢmay be appropriate probe to study the molecular mechanism of steroid saponins for antiplatelet activity.
We further investigated the characteristic of Timosaponin AⅢfor anti-platelet activity. Platelets react to a variety of biologically active substances (agonists) with partial or full activation characterized by shape change, release of granule contents, and aggregation. Conversely, other agents (antagonists) induce a state of reduced responsiveness to agonists. In our study, we demonstrated that Timosaponin AⅢinhibited the processes of platelet activation, such as platelet aggregation, platelet shape change, plateletαIIbβ3 activation, granule secretion and protein phosphorylation. Timosaponin AⅢinhibited the U46619-,arachidonic acid-, collagen-, and ADP-induced platelet aggregation in a concentration-dependent manner, but the sensitivity of platelet agonists varied to the inhibitory effect of Timosaponin AⅢ, U46619- and arachidonic acid -mediated platelet aggregation being more sensitive, suggesting that Timosaponin AⅢmay interfere with the TXA2 synthesis or its action directly. We observed an interesting phenomenon that Timosaponin AⅢby itself caused a shape change without aggregation. prostaglandin E1, A synthetic compound,inhibits platelets activity by stimulating the Gs-coupled prostacyclin IP receptor leading to the activation of adenylyl cyclase and production of cAMP. Cyclic AMP is a control molecule in platelets that interrupts multiple signaling pathways. Elevation cAMP levels can inhibit most platelet responses. However, Timosaponin AⅢdid not significantly increase the level cAMP in this study. Then, Timosaponin AⅢis also a potent platelet inhibitor as PGE1, but inhibits platelet activity independent of elevation cAMP levels of platelets.
The PKC family has long been know to involved in a number of platelet activity processes, most importantly aggregation and secretion. PMA can induce aggregation and secretion by activating PKC. Timosaponin AⅢdid not inhibited PMA–induced aggregation and secretion , suggesting Timosaponin AⅢmay inhibit the upstream signals of PKC. Thrombin interacts with platelet through a specific receptor belonging to the class or superfamily of receptors that are coupled to G proteins and phospholipase C, producing diacylglycerol, which stimulates protein kinase C that is closely linked to secretion. Activation of platelets by thrombin depends on TXA2 formation only at lower concentrations; at higher concentrations the ability of thrombin to activate platelets is independent of TXA2. In the present study, a higher concentration of thrombin was selected to induce platelet aggregation, and Timosaponin AⅢdidn’t inhibit thrombin-induced aggregation and secretion. The inhibitory effect of Timosaponin AⅢwas selective for U46619 that activated TXA2-mediated platelet aggregation.The ERKs are phosphorylated, and presumably activated, by various agonists such as thrombin, PMA and U46619, but their role and relevance in platelet function remain unclear. Our study further confirms ERK 1/2 activation in response to platelet stimulation with agonists. Moreover, we have demonstrated that Timosaponin AⅢis efficient inhibitors of ERK 1/2 phosphorylation induced by U46619. This inhibitory effect could involve blockage of signaling downstream TP, since these Timosaponin AⅢhave a minor effect on TXA2-independent thrombin-induced ERK1/2 phosphorylation and PMA-induced ERK phosphorylation.TXA2 is a metabolite of arachidonic acid with a chemical half-life of about 30 s. TXA2 released from activated platelets binds to TXA2 receptors and causes platelet shape change and aggregation as a positive feedback mediator. TXA2 receptor interacts with heterotrimeric G proteins Gq and G12/13. Gq-mediated phospholipase Cβactivation appears to play a central role in platelet aggregation and secretion, whereas G12/13-signaling pathway induces platelet shape change, involving Rho/Rho kinase-mediated phosphorylation of the myosin light chain. Our further research found Timosaponin AⅢselectively blockaded TP-mediated Gq-signaling pathway, and had no effect on G12/13-signaling pathway. We observed U73122, a potent inhibitor of PLCβ, inhibited Timosaponin AⅢ-induced shape change, suggesting Timosaponin AⅢbeing a partial agonist by activating Gq. Finally, we found that Timosaponin AⅢhas a synergistic action with SQ29548, a selective TXA2 receptor antagonist, on U46619-induced platelet aggregation.
In conclusion, our present study first used Timosaponin AⅢ, isolated from the root of Anemarrhenae asphodeloides Bge (Liliaceae) , as an ideal probe to explore the molecular mechanism of anti-platelet activity of steroid saponins.. Based on signal pathway, we supposed that candidate target of Timosaponin AⅢon anti-platelet activity might be TXA2 receptor .
甾体皂苷是一类重要的中药活性成分,在许多植物中都含有甾体皂苷成分。它具有广泛的药理作用和重要的生物活性,例如防治心血管疾病、抗肿瘤、抗真菌等作用。甾体皂苷的基本骨架是由含27个碳的苷元结构与一个或多个糖链连接构成的,主要分为含一个糖链(主要在C3-位)的螺甾皂苷(spirostanol saponins)和含2个糖链(主要在C 3-及C26-位)的呋甾皂苷(furostanol saponins)。甾体皂苷具有较高的医药价值,已有报道多种甾体皂苷具有明确的抗血小板活性作用,但目前对甾体皂苷抗血小板作用机制还了解很少。本研究的目的就是揭示甾体皂苷抗血小板活性的作用机制。
我们首先筛选了六个从知母中提取和合成的单体化合物,发现知母皂苷AⅢ抑制血小板聚集的作用最强。进一步构效关系分析显示,苷元结构和C-3位糖链组成和连接方式与甾体皂苷抑制血小板聚集活性密切相关。体内研究结果表明知母皂苷AⅢ延长小鼠尾静脉出血时间,保护致死性的血栓栓塞。这些结果提示知母皂苷AⅢ是研究甾体皂苷抗血小板活性的理想小分子探针。
我们进一步研究知母皂苷AⅢ抗血小板活性的作用特点。血小板激活包括变形、聚集、释放等反应。血小板抑制剂可以抑制血小板的激活反应。在我们研究中,知母皂苷AⅢ抑制了血小板聚集、变形、释放、αIIbβ3激活、蛋白磷酸化等激活反应。同时,知母皂苷AⅢ也表现出其抗血小板活性的自身特点,知母皂苷AⅢ虽然剂量依赖性地抑制U46619(TXA2受体激动剂)、AA、胶原和ADP诱导的血小板聚集,但对各诱导剂的敏感性不同,对U46619和AA的作用最强,这提示知母皂苷AⅢ可能阻断TXA2生成或直接作用TXA2通路上;观察到知母皂苷AⅢ本身也会引起血小板变形,而PGE1则没有观察到这一现象。PGE1与血小板膜上IP受体结合,激活Gs通路,升高细胞内cAMP。升高的cAMP能阻断多条血小板信号通路激活而抑制血小板的激活反应。然而,在我们的实验中并未观察到知母皂苷AⅢ升高血小板cAMP的作用。知母皂苷AⅢ与PGE1相似,是一个有效的血小板抑制剂,但其发挥作用并不是通过升高cAMP。
PKC的激活与血小板激活反应有关,主要调节血小板的聚集和释放。PMA激活PKC诱导血小板聚集和释放,然而,知母皂苷AⅢ并没有抑制PMA诱导的血小板变形和释放,这提示知母皂苷AⅢ可能抑制PKC上游信号通路。凝血酶与血小板膜G蛋白偶联受体结合,激活磷脂酶C,产生DAG,导致PKC的激活,引起血小板的释放反应。凝血酶激活血小板仅在低浓度作用时依赖TXA2的生成,而高浓度时不依赖TXA2的生成。使用高浓度凝血酶诱导血小板聚集,知母皂苷AⅢ不能抑制高浓度凝血酶诱导的血小板聚集,而选择性抑制TXA2受体激动剂U46619介导的血小板聚集和释放。我们的实验证实了U46619、凝血酶和PMA都能诱导血小板ERK的磷酸化,知母皂苷AⅢ能有效地抑制U46619诱导ERK1/2磷酸化,而对不依赖TXA2受体介导的凝血酶和PMA诱导的ERK1/2磷酸化没有作用,这表明知母皂苷AⅢ能阻断TXA2受体介导的信号通路的激活。血小板激活生成TXA2分泌后与血小板膜表面TXA2受体结合,而TXA2受体主要偶联在Gq和G12/13两条通路上,Gq通路介导的PLCβ激活在激动剂诱导的血小板聚集和释放中有至关重要的作用;G12/13介导的Rho/Rho激酶途径主要与血小板的变形有关。我们研究表明,知母皂苷AⅢ选择性阻断TXA2受体介导的Gq通路的激活,而不影响G12/13通路。此外,磷脂酶C抑制剂能抑制知母皂苷AⅢ诱导的血小板变形,知母皂苷AⅢ诱导的变形很可能由于部分激活了Gq通路。最后,我们发现知母皂苷AⅢ能协同TXA2受体拮抗剂SQ29548共同抑制U46619诱导的血小板聚集。
总的来说,我们选择知母皂苷AⅢ作为理想的小分子探针研究甾体皂苷抗血小板活性的作用机制,并在研究信号转导通路的基础上,推测TXA2受体可能是知母皂苷AⅢ抗血小板活性作用的候选靶点。
Platelets, anucleated cells that originate from bone marrow megakaryocytes, are essential for primary hemostasis in the blood. In addition, a role for platelets in pathogenesis of a number of cardiovascular events is recognized. Increasing evidence indicates that enhanced platelet activation plays a critical role in initiation and development of atherothrombotic diseases. Accordingly, to prevent and treat thrombosis and vascular diseases, antiplatelet therapy may be a potential strategy. Many antiplatelet agents such as aspirin (an inhibitor of cyclooxygenase (COX)), clopidogrel, and ticlopidine (adenosine diphosphate receptor antagonists acting at the purinergic receptors of P2Y), have been reported to be beneficial to patients suffering from cardiovascular disorders. However, some drawbacks of these drugs such as aspirin resistance, the risk of neutropenia, bleeding, and thrombocytopenia, were occasionally observed. Therefore, development of more safe and effective antiplatelet agents may be a promising approach for prevention and treatment of atherothrombosis.
Steroidal saponins are widely distributed in plants and have many pharmacologic actions and biological activities, such as antiplatelet activity, antitumor activity, and antidementia activity. They are an important class of natural products which is composed of a C-27 aglycone moiety and sugar chains of one or more monosaccharides. These compounds are classified as spirostanol glycosides with a sugar chain at C3 position, and furostanol saponins with two sugar chains at both C3 and C26 positions respectively. They have considerable potential as pharmaceutical and/or nutraceutical agents in natural or synthetic form. Steroidal glycosides, from a variety of sources, have been reported to have platelet inhibitory or aggregative activity, such as sarsasapogenin glycosides extracted from the rhizome of Anemarrhenae asphodeloides Bge (Liliaceae), has shown strong anti-platelet activity. But until now, little is known about molecular mechanism for the anti-platelet activity .Our aim is to determine the mechanism of steroidal saponins of anti-platelet activity.
We first screened 6 natural steroidal saponins isolated from the root of Anemarrhenae asphodeloides Bge (Liliaceae) for their effects on rat platelet aggregation. We found that Timosaponin AⅢexhibited stronger inhibitory effects on platelet aggregation than other compounds, Further structure–activity assay revealed that the activity of Timosaponin AⅢin inhibiting platelet aggregation was not only attributed in part to the number, the length and the type of sugar side chains attached by a glycoside at C-3, but also related with the aglycon part. In the in vivo study, Timosaponin AⅢprolonged mouse tail bleeding time using bleeding time model and protected mice against fatal thrombosis in a concentration dependent manner. These results indicated that Timosaponin AⅢmay be appropriate probe to study the molecular mechanism of steroid saponins for antiplatelet activity.
We further investigated the characteristic of Timosaponin AⅢfor anti-platelet activity. Platelets react to a variety of biologically active substances (agonists) with partial or full activation characterized by shape change, release of granule contents, and aggregation. Conversely, other agents (antagonists) induce a state of reduced responsiveness to agonists. In our study, we demonstrated that Timosaponin AⅢinhibited the processes of platelet activation, such as platelet aggregation, platelet shape change, plateletαIIbβ3 activation, granule secretion and protein phosphorylation. Timosaponin AⅢinhibited the U46619-,arachidonic acid-, collagen-, and ADP-induced platelet aggregation in a concentration-dependent manner, but the sensitivity of platelet agonists varied to the inhibitory effect of Timosaponin AⅢ, U46619- and arachidonic acid -mediated platelet aggregation being more sensitive, suggesting that Timosaponin AⅢmay interfere with the TXA2 synthesis or its action directly. We observed an interesting phenomenon that Timosaponin AⅢby itself caused a shape change without aggregation. prostaglandin E1, A synthetic compound,inhibits platelets activity by stimulating the Gs-coupled prostacyclin IP receptor leading to the activation of adenylyl cyclase and production of cAMP. Cyclic AMP is a control molecule in platelets that interrupts multiple signaling pathways. Elevation cAMP levels can inhibit most platelet responses. However, Timosaponin AⅢdid not significantly increase the level cAMP in this study. Then, Timosaponin AⅢis also a potent platelet inhibitor as PGE1, but inhibits platelet activity independent of elevation cAMP levels of platelets.
The PKC family has long been know to involved in a number of platelet activity processes, most importantly aggregation and secretion. PMA can induce aggregation and secretion by activating PKC. Timosaponin AⅢdid not inhibited PMA–induced aggregation and secretion , suggesting Timosaponin AⅢmay inhibit the upstream signals of PKC. Thrombin interacts with platelet through a specific receptor belonging to the class or superfamily of receptors that are coupled to G proteins and phospholipase C, producing diacylglycerol, which stimulates protein kinase C that is closely linked to secretion. Activation of platelets by thrombin depends on TXA2 formation only at lower concentrations; at higher concentrations the ability of thrombin to activate platelets is independent of TXA2. In the present study, a higher concentration of thrombin was selected to induce platelet aggregation, and Timosaponin AⅢdidn’t inhibit thrombin-induced aggregation and secretion. The inhibitory effect of Timosaponin AⅢwas selective for U46619 that activated TXA2-mediated platelet aggregation.The ERKs are phosphorylated, and presumably activated, by various agonists such as thrombin, PMA and U46619, but their role and relevance in platelet function remain unclear. Our study further confirms ERK 1/2 activation in response to platelet stimulation with agonists. Moreover, we have demonstrated that Timosaponin AⅢis efficient inhibitors of ERK 1/2 phosphorylation induced by U46619. This inhibitory effect could involve blockage of signaling downstream TP, since these Timosaponin AⅢhave a minor effect on TXA2-independent thrombin-induced ERK1/2 phosphorylation and PMA-induced ERK phosphorylation.TXA2 is a metabolite of arachidonic acid with a chemical half-life of about 30 s. TXA2 released from activated platelets binds to TXA2 receptors and causes platelet shape change and aggregation as a positive feedback mediator. TXA2 receptor interacts with heterotrimeric G proteins Gq and G12/13. Gq-mediated phospholipase Cβactivation appears to play a central role in platelet aggregation and secretion, whereas G12/13-signaling pathway induces platelet shape change, involving Rho/Rho kinase-mediated phosphorylation of the myosin light chain. Our further research found Timosaponin AⅢselectively blockaded TP-mediated Gq-signaling pathway, and had no effect on G12/13-signaling pathway. We observed U73122, a potent inhibitor of PLCβ, inhibited Timosaponin AⅢ-induced shape change, suggesting Timosaponin AⅢbeing a partial agonist by activating Gq. Finally, we found that Timosaponin AⅢhas a synergistic action with SQ29548, a selective TXA2 receptor antagonist, on U46619-induced platelet aggregation.
In conclusion, our present study first used Timosaponin AⅢ, isolated from the root of Anemarrhenae asphodeloides Bge (Liliaceae) , as an ideal probe to explore the molecular mechanism of anti-platelet activity of steroid saponins.. Based on signal pathway, we supposed that candidate target of Timosaponin AⅢon anti-platelet activity might be TXA2 receptor .
引文
1. Gü?lü-Ustünda? O, Mazza G. Saponins: properties, applications and processing [J]. Crit Rev Food Sci Nutr. 2007; 47(3):231-58.
2. Sparg SG, Light ME, Staden JV. Biological activities and distribution of plant saponins [J]. J Ethnopharmacol2004; 94: 219–43.
3. Rao AV, Gurfinkel DM. The bioactivity of saponins: triterpenoid and steroidal glycosides [J]. Drug Metabol Drug Interact 2000; 17: 211-35.
4. DavìG, Patrono C. Platelet activation and atherothrombosis [J]. N Engl J Med. 2007; 357(24):2482-94.
5. Offermanns S. Activation of platelet function through G protein-coupled receptors [J]. Circ Res. 2006; 99(12):1293-304.
6. Balamurugan R, Dekker FJ, Waldmann H. Design of compound libraries based on natural product scaffolds and protein structure similarity clustering (PSSC) [J]. Mol Biosyst. 2005; 1(1):36-45.
7. Bredel M & Jacoby E. Chemogenomics: an emerging strategy for rapid target and drug discovery [J]. Nat Rev Genet. 2004; 5(4):262-75.
8. Francis G, Kerem Z, Makkar HP, Becker K. The biological action of saponins in animal systems: a review [J]. Br J Nutr. 2002; 88 (6): 587-605
9.徐先祥,刘青云,彭代银.中药皂苷类成分抗血小板作用研究概况.中国中西医结合杂志[J]. 2001;21(2):150-152
10. Jianying Zhang, Zhiyun Meng, Meiying Zhang, Deshun Ma, Sioxi Xu, Hiroyuki Kodama. Effect of six steroidal saponins isolated from anemarrhenae rhizoma on platelet aggregation and hemolysis in human blood [J]. Clinica Chimica Acta. 1999;289:79-88
11.赵泰.现代中草药成分化学[M].北京:中国医药科技出版社,2002.
12. Nakashima N, Kimura I, Kimura M et al. Isolation of pseudoprototimosaponin AⅢrhizomes of Anemarrhena asphodeloides and its hypoglycemic activity in streptozotocin-induced diabetic mice [J]. J Nat Prod. 1993, 56(3): 345-350
13. Niwa A, Takeda O, Ishimaru M et al. Screening test for platelet aggregation inhibitor in natural products. The active principle of Anemarrhenae Rhizoma [J]. Yakugaku Zasshi, 1988,108(6):555–561
14.康利平,马百平,史天军等.知母中的两种新呋甾皂苷.药学学报[J]. 2006,42(6):527-532
15. Frank W, King, Sylvia Fong, et al. Timosaponin AIII is preferentially cytotoxic to tumor cells through inhibition of mTOR and induction of ER Stress [J].Plosone, 2009, 4(9):1-12
16. GrorgeJ, Broze, Jr,Zheng-FengYin,Nina Lasky et al.A tail vein bleeding time model and delayed bleeding in hemophiliac mice[J]. Thromb Haemost 2001,(85):747-748
17. Guei-Jane Wang, Lie-Chwen Lin, Chieh-Fu Chen et al. Effect of timosaponin A-III, from Anemarrhenae asphodeloides Bunge (Liliaceae), on calcium mobilization in vascular endothelial[J]. Life Sciences.2002,(71):1081–1090
18. Bomi Lee, Kangsik Jung, Dong-Hyun Kim. Timosaponin AIII, a saponin isolated from Anemarrhena asphodeloides, ameliorates learning and memory deficits in mice [J]. Pharmacology, Biochemistry and Behavior.2009,(93):121-127
19. Crittenden JR, Bergmeier W, Zhang Y, Piffath CL, Liang Y, Wagner DD, Housman DE, Graybiel AM. CalDAG-GEFI integrates signaling for platelet aggregation and thrombus formation [J]. Nat Med. 2004; 10(9):982-6.
20. Nishizuka, Y. 1992. Intracellular signaling by hydrolysis of phospholipids andactivation of protein kinase C [J]. Science 258: 607–614.
21. Stitham J, Arehart EJ, Gleim SR, Douville KL, Hwa J. Human prostacyclin receptor structure and function from naturally-occurring and synthetic mutations[J]. Prostaglandins Other Lipid Mediat. 2007; 82(1-4):95-108.
22. Offermanns S. Activation of platelet function through G protein-coupled receptors [J]. Circ Res 2006; (99):1293-304.
23. Coughlin SR. Protease-activated receptors in hemostasis, thrombosis and vascular biology [J]. J Thromb Haemost. 2005; (3):1800–1814.
24. Kobayashi T, Tahara Y, Matsumoto M, Iguchi M, Sano H, Murayama T,Arai H,Oida H, Yurugi-Kobayashi T, Yamashita JK, Katagiri H, Majima M, Yokode M, Kita T, Narumiya S. Roles of thromboxane A(2) and prostacyclin in the development of atherosclerosis in apoE-deficient mice[J]. J Clin Invest. 2004;(114):784–794
25. Li Z, Xi X, Du X. A mitogen-activated protein kinase– dependent signaling pathway in the activation of platelet integrin alpha IIb beta3 [J]. J Biol Chem. 2001; (276):42226-42232.
26. Wu Y, Asazuma N, Satoh K, et al. Interaction between von Willebrand factor and glycoprotein Ib activates Src kinase in human platelets: role of phosphoinositide 3-kinase [J]. Blood. 2003; (101): 3469-3476.
27. Mangin P, Yuan Y, Goncalves I, et al. Signaling role for phospholipase C gamma 2 in platelet glycoprotein Ib alpha calcium flux and cytoskeletal reorganization: involvement of a pathway distinct from FcR gamma chain and Fc gamma RIIA[J]. J Biol Chem. 2003;(278):32880-32891
28. Kobayashi T, Tahara Y, Matsumoto M, Iguchi M, Sano H, Murayama T,Arai H, Oida H, Yurugi-Kobayashi T, Yamashita JK, Katagiri H, Majima M, Yokode M, Kita T, Narumiya S. Roles of thromboxane A(2) and prostacyclin in the development of atherosclerosis in apoE-deficient mice[J]. J Clin Invest. 2004;(114):784–794
29. Watson SP. Collagen receptor signaling in platelets and megakaryocytes [J]. Thromb Haemost. 1999; (82):365-376.
30. Moon J. Cho, Junling Liu, Tamara I. Pestina, et al. The roles ofαIIbβ3-mediated outside-in signal transduction, thromboxane A2, and adenosine diphosphate in collagen-induced platelet aggregation [J]. Hemostasis, thrombosis and varscularbiology. 2003;(10)1:2646-2652
31. Satoko Ohkubo, Norimichi Nakahata , Yasushi Ohizumi. Thromboxane A2-mediated shape change: independent of Gqphospholipase C- Ca2+ pathway in rabbit platelets [J]. British Journal of Pharmacology. 1996;(117): 1095 -1104
32.罗国安、梁琼麟、张荣利、王义明、刘清飞、胡坪.化学物质组学与中药方剂研究--兼析清开灵复方物质基础研究[J].世界科学技术-中医药现代化. 2006;8(1):6-15.
33. Foster CJ, Prosser DM, Agans JM, et al. Molecular identification and characterization of the platelet ADP receptor targeted by thienopyridine antithrombotic drugs [J]. J Clin Invest. 2001; (107):1591-1598
34. Haslam RJ, Dickinson NT, Jang EK. Cyclic nucleotides and phosphodiesterases in platelets [J]. Thromb Haemost. 1999;(82):412-423
35. Bredel M & Jacoby E. Chemogenomics: an emerging strategy for rapid target and drug discovery [J]. Nat Rev Genet. 2004; 5(4):262-275.
36. Buensuceso CS, Obergfell A, Soriani A, et al. Regulation of outside-in signaling in platelets by integrin-associated protein kinase C beta [J]. J Biol Chem. 2005(280):644-653.
37. Coughlin SR. Protease-activated receptors in hemostasis, thrombosis and vascular biology [J]. J Thromb Haemost. 2005;(3):1800–1814
38. Kahn ML, Zheng YW, Huang W, Bigornia V, Zeng D, Moff S, Farese RV Jr, Tam C, Coughlin SR. A dual thrombin receptor system for platelet activation [J]. Nature. 1998;(394):690–694
39. Kahn ML, Nakanishi-Matsui M, Shapiro MJ, Ishihara H, Coughlin SR. Protease-activated receptors 1 and 4 mediate activation of human platelets by thrombin[J]. J Clin Invest. 1999; (103):879–887.
40. Djellas Y, Manganello JM, Antonakis K, Le Breton GC. Identification of Galpha13 as one of the G-proteins that couple to human platelet thromboxane A2 receptors[J]. J Biol Chem. 1999; (274):14325–14330.
41. Thomas DW, Mannon RB, Mannon PJ, Latour A, Oliver JA, Hoffman M, Smithies O, Koller BH, Coffman TM. Coagulation defects and altered hemodynamic responses in mice lacking receptors for thromboxane A2 [J]. J Clin Invest. 1998;(102):1994–2001
42. Yang J, Wu J, Kowalska MA, Dalvi A, Prevost N, O’Brien PJ, Manning D, Poncz M, Lucki I, Blendy JA, Brass LF. Loss of signaling through the G protein, Gz, results in abnormal platelet activation and altered responses to psychoactive drugs [J]. Proc Natl Acad Sci U S A. 2000;(97):9984–9989
43. Siess W, Lapetina EG. Ca2_ mobilization primes protein kinase C in human
44. Gilligan DM, Sarid R, Weese J. Adducin in platelets: activation-induced phosphorylation by PKC and proteolysis by calpain [J]. Blood. 2002; (99):2418-2426.
45. Gabbeta J, Yang X, Kowalska MA, Sun L, Dhanasekaran N, Rao AK. Platelet signal transduction defect with Galpha subunit dysfunction and diminished Galphaq in a patient with abnormal platelet responses [J]. Proceedings of the National Academy of Sciences of the United States of America 1997; (94):8750–87555.
46. Walker TR, Watson SP. Synergy between Ca2+ and protein kinase C is the major factor in determining the level of secretion from human platelets [J]. The Biochemical Journal 1993;289 (Pt 1):277–282.
47. Bernhard Nieswandt, Valerie Schult, Alexandra Zywietz, Costimulation of Gi- and G12/G13-mediated signaling pathways induces integrinαIIbβ3 activation in platelets [J]. J Biol Chem.2002;(207):9493–39498
48. Dorsam, R. T., Kim, S., Jin, J. and Kunapuli, S. P. (2002) Coordinated signaling through both G12/13 and Gi pathways is sufficient to activate GPIIb/IIIa in human platelets[J]. J. Biol. Chem. 277, 47588–47595
49. Nieswandt, B., Schulte, V., Zywietz, A., Gratacap, M. P. and Offermanns, S. (20 Costimulation of Gi- and G12/G13-mediated signaling pathways induces integrinαIIbβ3 activation in platelets [J]. J. Biol. Chem. 2002;(277):39493–39498
50. Klages, B., Brandt, U., Simon, M. I., Schultz, G et al.. Activation of G12/G13 results in shape change and Rho/Rho-kinase-mediated myosin light chain phosphorylation in mouse platelets [J]. J. Cell Biol.1999;(144); 745–754
51. Paul, B. Z., Daniel, J. L. and Kunapuli, S. P. Platelet shape change is mediated by both calcium-dependent and -independent signaling pathways. Role of p160 Rho-associated coiled-coil-containing protein kinase in platelet shape change [J]. J. Biol. Chem.1999;( 274): 28293–28300
52. Bauer, M., Retzer, M., Wilde, J. I., Maschberger, P., Essler, M., A et al. Dichotomous regulation of myosin phosphorylation and shape change by 50Rho-kinase and calcium in intact human platelets [J]. Blood.1999; (94):1665–1672
1 . Ruggeri ZM. Platelets in atherothrombosis. Nat Med. 2002;8:1227–1234.
2 .Fitzgerald GA. Mechanisms of platelets activation:thromboxaneA2 as an amplifying signal for other agonists. Am J Cardiol.1991;68:11B–15B.
3.Gachet C. ADP receptors of platelets and their inhibition. Thromb Haemost. 2001;86:222–232.
4.Kam PCA, Nethery CM. The thienopyridine derivatives (platelet adenosine diphosphate receptor antagonists), pharmacology and clinical developments. Anaesthesia. 2003;58:28–35.
5 .Casserly IP, Topol EJ. Glycoprotein IIb/IIIa antagonists—from bench to practice. Cell Mol Life Sci. 2002; 59:478–500.
6.Hodivala-Dilke KM, McHugh KP, Tsakiris DA, Rayburn H, Crowley D, Ullman-Cullere M et al. . Beta3-integrin-deficient mice are a model for Glanzmann thrombasthenia showing placental defects and reduced survival. J Clin Invest.1999; 103: 229–238.
7.Wu DM, Meiring M, Kotze HF, Deckmyn H, Cauwenberghs N (2002). Inhibition of platelet glycoprotein Ib, glycoprotein IIb/IIIa, or both by monoclonal antibodies prevents arterial thrombosis in baboons. Arterioscler Thromb Vasc Biol 22: 323–328.
8 .Holtkotter O, Nieswandt B, Smyth N, Muller W, Hafner M, Schulte V et al. (2002). Integrin alpha 2-deficient mice develop normally, are fertile, but display partially defective platelet interaction with collagen. J Biol Chem 277: 10789–10794.
9.Kato K, Kanaji T, Russell S, Kunicki TJ, Furihata K, Kanaji S et al. (2003). The contribution of glycoprotein VI to stable platelet adhesion and thrombus formation illustrated by targeted gene deletion. Blood 102: 1701–170
10 .Mangin P, Yap CL, Nonne C, Sturgeon SA, Goncalves I, Yuan Y et al. (2006).Thrombin overcomes the thrombosis defect associated with platelet GPVI/FcR gamma deficiency. Blood 107: 4346–4353
11. Watanabe N, Nakajima H, Suzuki H, Oda A, Matsubara Y, Moroi M et al. (2003). Functional phenotype of phosphoinositide 3-kinase p85alpha-null platelets characterized by an impaired response to GP VI stimulation. Blood 102: 541–548.
12 .Jackson SP, Schoenwaelder SM, Goncalves I, Nesbitt WS, Yap CL, Wright CE et al. . PI 3-kinase p110beta: a new target for antithrombotic therapy. Nat Med .2005;11: 507–514
13.史秀丽,傅佳,李光武1血小板靶向中药有效成分及单体研究进展[J ]1中国中药杂志, 2006, 31 (5) : 36123651
2. Sparg SG, Light ME, Staden JV. Biological activities and distribution of plant saponins [J]. J Ethnopharmacol2004; 94: 219–43.
3. Rao AV, Gurfinkel DM. The bioactivity of saponins: triterpenoid and steroidal glycosides [J]. Drug Metabol Drug Interact 2000; 17: 211-35.
4. DavìG, Patrono C. Platelet activation and atherothrombosis [J]. N Engl J Med. 2007; 357(24):2482-94.
5. Offermanns S. Activation of platelet function through G protein-coupled receptors [J]. Circ Res. 2006; 99(12):1293-304.
6. Balamurugan R, Dekker FJ, Waldmann H. Design of compound libraries based on natural product scaffolds and protein structure similarity clustering (PSSC) [J]. Mol Biosyst. 2005; 1(1):36-45.
7. Bredel M & Jacoby E. Chemogenomics: an emerging strategy for rapid target and drug discovery [J]. Nat Rev Genet. 2004; 5(4):262-75.
8. Francis G, Kerem Z, Makkar HP, Becker K. The biological action of saponins in animal systems: a review [J]. Br J Nutr. 2002; 88 (6): 587-605
9.徐先祥,刘青云,彭代银.中药皂苷类成分抗血小板作用研究概况.中国中西医结合杂志[J]. 2001;21(2):150-152
10. Jianying Zhang, Zhiyun Meng, Meiying Zhang, Deshun Ma, Sioxi Xu, Hiroyuki Kodama. Effect of six steroidal saponins isolated from anemarrhenae rhizoma on platelet aggregation and hemolysis in human blood [J]. Clinica Chimica Acta. 1999;289:79-88
11.赵泰.现代中草药成分化学[M].北京:中国医药科技出版社,2002.
12. Nakashima N, Kimura I, Kimura M et al. Isolation of pseudoprototimosaponin AⅢrhizomes of Anemarrhena asphodeloides and its hypoglycemic activity in streptozotocin-induced diabetic mice [J]. J Nat Prod. 1993, 56(3): 345-350
13. Niwa A, Takeda O, Ishimaru M et al. Screening test for platelet aggregation inhibitor in natural products. The active principle of Anemarrhenae Rhizoma [J]. Yakugaku Zasshi, 1988,108(6):555–561
14.康利平,马百平,史天军等.知母中的两种新呋甾皂苷.药学学报[J]. 2006,42(6):527-532
15. Frank W, King, Sylvia Fong, et al. Timosaponin AIII is preferentially cytotoxic to tumor cells through inhibition of mTOR and induction of ER Stress [J].Plosone, 2009, 4(9):1-12
16. GrorgeJ, Broze, Jr,Zheng-FengYin,Nina Lasky et al.A tail vein bleeding time model and delayed bleeding in hemophiliac mice[J]. Thromb Haemost 2001,(85):747-748
17. Guei-Jane Wang, Lie-Chwen Lin, Chieh-Fu Chen et al. Effect of timosaponin A-III, from Anemarrhenae asphodeloides Bunge (Liliaceae), on calcium mobilization in vascular endothelial[J]. Life Sciences.2002,(71):1081–1090
18. Bomi Lee, Kangsik Jung, Dong-Hyun Kim. Timosaponin AIII, a saponin isolated from Anemarrhena asphodeloides, ameliorates learning and memory deficits in mice [J]. Pharmacology, Biochemistry and Behavior.2009,(93):121-127
19. Crittenden JR, Bergmeier W, Zhang Y, Piffath CL, Liang Y, Wagner DD, Housman DE, Graybiel AM. CalDAG-GEFI integrates signaling for platelet aggregation and thrombus formation [J]. Nat Med. 2004; 10(9):982-6.
20. Nishizuka, Y. 1992. Intracellular signaling by hydrolysis of phospholipids andactivation of protein kinase C [J]. Science 258: 607–614.
21. Stitham J, Arehart EJ, Gleim SR, Douville KL, Hwa J. Human prostacyclin receptor structure and function from naturally-occurring and synthetic mutations[J]. Prostaglandins Other Lipid Mediat. 2007; 82(1-4):95-108.
22. Offermanns S. Activation of platelet function through G protein-coupled receptors [J]. Circ Res 2006; (99):1293-304.
23. Coughlin SR. Protease-activated receptors in hemostasis, thrombosis and vascular biology [J]. J Thromb Haemost. 2005; (3):1800–1814.
24. Kobayashi T, Tahara Y, Matsumoto M, Iguchi M, Sano H, Murayama T,Arai H,Oida H, Yurugi-Kobayashi T, Yamashita JK, Katagiri H, Majima M, Yokode M, Kita T, Narumiya S. Roles of thromboxane A(2) and prostacyclin in the development of atherosclerosis in apoE-deficient mice[J]. J Clin Invest. 2004;(114):784–794
25. Li Z, Xi X, Du X. A mitogen-activated protein kinase– dependent signaling pathway in the activation of platelet integrin alpha IIb beta3 [J]. J Biol Chem. 2001; (276):42226-42232.
26. Wu Y, Asazuma N, Satoh K, et al. Interaction between von Willebrand factor and glycoprotein Ib activates Src kinase in human platelets: role of phosphoinositide 3-kinase [J]. Blood. 2003; (101): 3469-3476.
27. Mangin P, Yuan Y, Goncalves I, et al. Signaling role for phospholipase C gamma 2 in platelet glycoprotein Ib alpha calcium flux and cytoskeletal reorganization: involvement of a pathway distinct from FcR gamma chain and Fc gamma RIIA[J]. J Biol Chem. 2003;(278):32880-32891
28. Kobayashi T, Tahara Y, Matsumoto M, Iguchi M, Sano H, Murayama T,Arai H, Oida H, Yurugi-Kobayashi T, Yamashita JK, Katagiri H, Majima M, Yokode M, Kita T, Narumiya S. Roles of thromboxane A(2) and prostacyclin in the development of atherosclerosis in apoE-deficient mice[J]. J Clin Invest. 2004;(114):784–794
29. Watson SP. Collagen receptor signaling in platelets and megakaryocytes [J]. Thromb Haemost. 1999; (82):365-376.
30. Moon J. Cho, Junling Liu, Tamara I. Pestina, et al. The roles ofαIIbβ3-mediated outside-in signal transduction, thromboxane A2, and adenosine diphosphate in collagen-induced platelet aggregation [J]. Hemostasis, thrombosis and varscularbiology. 2003;(10)1:2646-2652
31. Satoko Ohkubo, Norimichi Nakahata , Yasushi Ohizumi. Thromboxane A2-mediated shape change: independent of Gqphospholipase C- Ca2+ pathway in rabbit platelets [J]. British Journal of Pharmacology. 1996;(117): 1095 -1104
32.罗国安、梁琼麟、张荣利、王义明、刘清飞、胡坪.化学物质组学与中药方剂研究--兼析清开灵复方物质基础研究[J].世界科学技术-中医药现代化. 2006;8(1):6-15.
33. Foster CJ, Prosser DM, Agans JM, et al. Molecular identification and characterization of the platelet ADP receptor targeted by thienopyridine antithrombotic drugs [J]. J Clin Invest. 2001; (107):1591-1598
34. Haslam RJ, Dickinson NT, Jang EK. Cyclic nucleotides and phosphodiesterases in platelets [J]. Thromb Haemost. 1999;(82):412-423
35. Bredel M & Jacoby E. Chemogenomics: an emerging strategy for rapid target and drug discovery [J]. Nat Rev Genet. 2004; 5(4):262-275.
36. Buensuceso CS, Obergfell A, Soriani A, et al. Regulation of outside-in signaling in platelets by integrin-associated protein kinase C beta [J]. J Biol Chem. 2005(280):644-653.
37. Coughlin SR. Protease-activated receptors in hemostasis, thrombosis and vascular biology [J]. J Thromb Haemost. 2005;(3):1800–1814
38. Kahn ML, Zheng YW, Huang W, Bigornia V, Zeng D, Moff S, Farese RV Jr, Tam C, Coughlin SR. A dual thrombin receptor system for platelet activation [J]. Nature. 1998;(394):690–694
39. Kahn ML, Nakanishi-Matsui M, Shapiro MJ, Ishihara H, Coughlin SR. Protease-activated receptors 1 and 4 mediate activation of human platelets by thrombin[J]. J Clin Invest. 1999; (103):879–887.
40. Djellas Y, Manganello JM, Antonakis K, Le Breton GC. Identification of Galpha13 as one of the G-proteins that couple to human platelet thromboxane A2 receptors[J]. J Biol Chem. 1999; (274):14325–14330.
41. Thomas DW, Mannon RB, Mannon PJ, Latour A, Oliver JA, Hoffman M, Smithies O, Koller BH, Coffman TM. Coagulation defects and altered hemodynamic responses in mice lacking receptors for thromboxane A2 [J]. J Clin Invest. 1998;(102):1994–2001
42. Yang J, Wu J, Kowalska MA, Dalvi A, Prevost N, O’Brien PJ, Manning D, Poncz M, Lucki I, Blendy JA, Brass LF. Loss of signaling through the G protein, Gz, results in abnormal platelet activation and altered responses to psychoactive drugs [J]. Proc Natl Acad Sci U S A. 2000;(97):9984–9989
43. Siess W, Lapetina EG. Ca2_ mobilization primes protein kinase C in human
44. Gilligan DM, Sarid R, Weese J. Adducin in platelets: activation-induced phosphorylation by PKC and proteolysis by calpain [J]. Blood. 2002; (99):2418-2426.
45. Gabbeta J, Yang X, Kowalska MA, Sun L, Dhanasekaran N, Rao AK. Platelet signal transduction defect with Galpha subunit dysfunction and diminished Galphaq in a patient with abnormal platelet responses [J]. Proceedings of the National Academy of Sciences of the United States of America 1997; (94):8750–87555.
46. Walker TR, Watson SP. Synergy between Ca2+ and protein kinase C is the major factor in determining the level of secretion from human platelets [J]. The Biochemical Journal 1993;289 (Pt 1):277–282.
47. Bernhard Nieswandt, Valerie Schult, Alexandra Zywietz, Costimulation of Gi- and G12/G13-mediated signaling pathways induces integrinαIIbβ3 activation in platelets [J]. J Biol Chem.2002;(207):9493–39498
48. Dorsam, R. T., Kim, S., Jin, J. and Kunapuli, S. P. (2002) Coordinated signaling through both G12/13 and Gi pathways is sufficient to activate GPIIb/IIIa in human platelets[J]. J. Biol. Chem. 277, 47588–47595
49. Nieswandt, B., Schulte, V., Zywietz, A., Gratacap, M. P. and Offermanns, S. (20 Costimulation of Gi- and G12/G13-mediated signaling pathways induces integrinαIIbβ3 activation in platelets [J]. J. Biol. Chem. 2002;(277):39493–39498
50. Klages, B., Brandt, U., Simon, M. I., Schultz, G et al.. Activation of G12/G13 results in shape change and Rho/Rho-kinase-mediated myosin light chain phosphorylation in mouse platelets [J]. J. Cell Biol.1999;(144); 745–754
51. Paul, B. Z., Daniel, J. L. and Kunapuli, S. P. Platelet shape change is mediated by both calcium-dependent and -independent signaling pathways. Role of p160 Rho-associated coiled-coil-containing protein kinase in platelet shape change [J]. J. Biol. Chem.1999;( 274): 28293–28300
52. Bauer, M., Retzer, M., Wilde, J. I., Maschberger, P., Essler, M., A et al. Dichotomous regulation of myosin phosphorylation and shape change by 50Rho-kinase and calcium in intact human platelets [J]. Blood.1999; (94):1665–1672
1 . Ruggeri ZM. Platelets in atherothrombosis. Nat Med. 2002;8:1227–1234.
2 .Fitzgerald GA. Mechanisms of platelets activation:thromboxaneA2 as an amplifying signal for other agonists. Am J Cardiol.1991;68:11B–15B.
3.Gachet C. ADP receptors of platelets and their inhibition. Thromb Haemost. 2001;86:222–232.
4.Kam PCA, Nethery CM. The thienopyridine derivatives (platelet adenosine diphosphate receptor antagonists), pharmacology and clinical developments. Anaesthesia. 2003;58:28–35.
5 .Casserly IP, Topol EJ. Glycoprotein IIb/IIIa antagonists—from bench to practice. Cell Mol Life Sci. 2002; 59:478–500.
6.Hodivala-Dilke KM, McHugh KP, Tsakiris DA, Rayburn H, Crowley D, Ullman-Cullere M et al. . Beta3-integrin-deficient mice are a model for Glanzmann thrombasthenia showing placental defects and reduced survival. J Clin Invest.1999; 103: 229–238.
7.Wu DM, Meiring M, Kotze HF, Deckmyn H, Cauwenberghs N (2002). Inhibition of platelet glycoprotein Ib, glycoprotein IIb/IIIa, or both by monoclonal antibodies prevents arterial thrombosis in baboons. Arterioscler Thromb Vasc Biol 22: 323–328.
8 .Holtkotter O, Nieswandt B, Smyth N, Muller W, Hafner M, Schulte V et al. (2002). Integrin alpha 2-deficient mice develop normally, are fertile, but display partially defective platelet interaction with collagen. J Biol Chem 277: 10789–10794.
9.Kato K, Kanaji T, Russell S, Kunicki TJ, Furihata K, Kanaji S et al. (2003). The contribution of glycoprotein VI to stable platelet adhesion and thrombus formation illustrated by targeted gene deletion. Blood 102: 1701–170
10 .Mangin P, Yap CL, Nonne C, Sturgeon SA, Goncalves I, Yuan Y et al. (2006).Thrombin overcomes the thrombosis defect associated with platelet GPVI/FcR gamma deficiency. Blood 107: 4346–4353
11. Watanabe N, Nakajima H, Suzuki H, Oda A, Matsubara Y, Moroi M et al. (2003). Functional phenotype of phosphoinositide 3-kinase p85alpha-null platelets characterized by an impaired response to GP VI stimulation. Blood 102: 541–548.
12 .Jackson SP, Schoenwaelder SM, Goncalves I, Nesbitt WS, Yap CL, Wright CE et al. . PI 3-kinase p110beta: a new target for antithrombotic therapy. Nat Med .2005;11: 507–514
13.史秀丽,傅佳,李光武1血小板靶向中药有效成分及单体研究进展[J ]1中国中药杂志, 2006, 31 (5) : 36123651
© 2004-2018 中国地质图书馆版权所有 京ICP备05064691号 京公网安备11010802017129号 地址：北京市海淀区学院路29号 邮编：100083 电话：办公室：(+86 10)66554848；文献借阅、咨询服务、科技查新：66554700 |